1. Field of the Invention
The present invention relates to processes and apparati for inactivating micro-organisms from liquids and improving the functional properties of food products using high pressure.
2. Background Art
Conventional thermal processing of milk, fruit juices and other foods to eliminate pathogens and control spoilage organisms may have adverse effects on flavor, vitamin content and functional properties or not completely inactivate undesirable enzymes such as pectinesterase (which causes the loss of cloud stability in fruit juices). Thus, there is a need to develop processing alternatives to conventional thermal processing for food production. In addition, foodborne illness due to the survival of pathogens in foods which were once considered safe due to intrinsic factors such as acidity (e.g., the October, 1996 Escherichia coli O157:H7 outbreak in Odwalla brand apple juice) may demand alternative processes which ensure food safety while protecting the delicate flavors of these foods. Additionally, there is a need for a process to sterilize heat-sensitive pharmaceutical preparations.
Physical phenomena such as microwaves, infrared, ultraviolet or gamma irradiation, high intensity laser or non-coherent light pulses, ultrasound, ohmic heating, pulsed electric fields, high voltage electric discharges ("electroporation"), bactericidal enzymes, mild heat with slight pressurization ("manothermosonication"), and extrusion cooking have been investigated singly or in combination as a means of processing dairy and other foods (Gallman and Eberhard, 1993, Gould, 1995, Knorr et al., 1994, Mertens and Knorr, 1992). Although additive or synergistic combinations of these technologies have been suggested as potential alternatives to conventional thermal processing which should improve food quality, functionality and safety, to date all these alternative methods have required the use of conventional heat exchangers to provide a sufficient microbicidal effect of these processes.
Nonthermal processing of foods, such as high hydrostatic pressure ("HHP") and high pressure homogenization ("HPH"), are considered a more "gentle" means of processing than conventional thermal processing. They will denature proteins, inactivate enzymes, gelatinize starches and inactivate microorganisms, while minimizing the loss of colors, flavors, aromas and vitamins and other nutrients. In addition, since these processes are non-thermal, foods so processed do not develop toxic components, and off-flavors are not produced. Another secondary advantage of non thermal processing by HHP and HPH is that gelled egg and seafood proteins are superior in many aspects to the gels formed by thermal processing.
High hydrostatic pressure (HHP)--the use of a high isostatic pressure to inactivate microbes and alter the physicochemical and functional properties of foods (e.g., form gels)--is currently in use commercially in a wide range of foods. Milk, juices and other liquid foods may be processed semi-continuously in a series of batched high-pressure vessels. Non-liquid foods are processed in packages of flexible films or other material stacked in a high-pressure vessel, and typically water is used as the carrier medium (Lehmann, 1996; Pothakamury et al., 1995). Pressure may be applied directly by high-pressure vessels of the piston-and-cylinder design or indirectly by pressure intensifiers attached to the vessels. Pressures vary depending on the application, but are ordinarily in the range of 400 to 900 MPa. In Japan, HHP is being used commercially on a wide variety of food products to inactivate microbes, produce excellent gels and retain more vitamins, natural flavors, colors and aromas than would be possible with thermal processing. Lehmann (1996) has reported that it should be possible to use the HHP technology to sterilize cosmetics and pharmaceuticals. Although HHP can be effective for certain applications, the technology has the disadvantages of being a batch process with long process times of an hour or more required to achieve desired results. Additionally, HHP has not proved reliable in terms of the degree of microbial inactivation accomplished under standard operating conditions.
HHP has been evaluated for a number of antimicrobial applications, ranging from the human-immunodeficiency virus that causes AIDS (Nakagami et al., 1996) to the herpes simplex virus (Nakagami et al., 1992) to a host of foodborne pathogens, "model pathogens" and spoilage organisms in a variety of food medi. In general, gram-negative bacteria arc most sensitive to HHP, followed by yeasts, complex viruses, molds, and gram-positive bacteria, but researchers have noted that there is much inter- and intra-species variation in the resistance of microorganisms to HHP (Cheftel, 1995) and that exponential-phase cells are much more resistant to HHP than stationary-phase cells (Cheftel, 1995, Mackey et al., 1995). Patterson et al. (1995) found that 15 min of HHP treatments of 350 MPa, 375 MPa, 450 MPa, 700 MPa, and 700 MPa were required to achieve 5 log cycle reductions in S. typhimurium, L. monocytogenes, S. enteritidis, E. coli O157:H7, and S. aureus respectively, and that there was significant variation in the baroresistance between different strains of both L. monocytogenes and E. coli O157:H7.
Additionally, the physicochemical composition of the medium may confer resistance to HHP (Cheftel, 1995, Mackey et al., 1995), and in general, microorganisms exhibit more resistance to HHP in certain foods than in buffers (Cheftel, 1995, Gervilla et al., 1997, Patterson et al., 1995).
Microbial inactivation by HHP may be improved by increasing processing temperatures to above 50.degree. C. or lowering to between -30 and 5.degree. C. (Cheftel, 1995). Still, most surviving cells are sublethally-injured and survival rates are dependent on a recovery period, which may overstate HHP effectiveness per se and indicates a need to combine this process with other technologies to achieve optimal microbial inactivation.
Thus, the disadvantages of HHP processing can be divided into at least three categories: economic feasibility, engineering limitations, and microbial safety concerns. The economic feasibility of commercial HHP is limited by the high cost of capital investment for new equipment (Mertens and Deplace, 1993) and the low productivity and high labor cost of the batch process. Some "semi-continuous" systems have been developed for the processing of juices and other liquid foods (Pothakamury et al., 1995), but these are only marginal improvements, in terms of economics. In the case of foods processed as discrete retail packages, there are the concerns of volume inefficiency, where space is wasted as package and inter-package volume, and inefficient process cycle times, which must include time for loading, unloading, pressurizing and decompressing. Feasibility is further limited by the long process times of 30 minutes to 1 hour required by some applications. While these feasibility problems may be somewhat mitigated by increasing the size of the pressure vessel, size is limited by engineering concerns, which call for thicker walls to accommodate even modest increases in internal volume. Current technology limits the size of pressure vessels that can be forged to an internal volume of approximately 700 L for an operating pressure of 500 MPa (Mertens and Deplace, 1993).
A critical problem with HHP processing is ensuring the effectiveness of microbial inactivation. Bacterial spores can survive HHP in excess of 1000 MPa, and some bacterial spores may be "superdormant" and not respond to conditions conducive to germination before HHP processing (Cheftel, 1995). Anaerobic spore-forming pathogens such as Clostridium botulinum and Clostridium perfrigens and spoilage organisms such as Bacillus stearothermophilus are a concern in low-acid foods preserved by high-temperature processing (Jay, 1992), and may present a serious problem in applying HHP processing to certain foods. In a review article, Hayakawa (1996) addressed this problem and demonstrated that repeated isostatic pressurization, i.e. 5-6 cycles of 5 minutes or longer duration during which the maximal pressure reaches 600 MPa and the temperature of the fluid is at 70.degree. C., can destroy Bacillus stearothermophilus spores, but this remains a harsh treatment for many fluids, and the requirement for cycling is impractical for most commercial applications. In addition, recent work indicates that microorganisms may be induced to develop resistance to HHP. Hauben et al. (1997) used alternating cycles of HHP treatment followed by outgrowth and cultivation of survivors to isolate three barorcsistant mutants of E. coli MG1655 which were able to survive HHP at 800 MPa for 15 min.
The use of HPH has been more limited than HHP, with applications being primarily the stabilization of emulsions and the extraction of intracellular enzymes and other proteins from plant and animal cells. Some investigation of the application of HPH to microbial inactivation has occurred in studies on the recovery of intracellular materials from yeasts and other microbial cultures, but the use of HPH in these studies was not to produce microbially-stable foods (Bailey et al., 1995, Baldwin and Robinson, 1992, Middelberg, 1995, Siddiqi et al., 1995, Siddiqi and Titchenerhooker, 1996). To liberate the desired intracellular products, cell disruption is achieved by multiple passes through the homogenizer at pressures ranging from approximately 10 to 100 MPa (Baldwin and Robinson, 1992, Siddiqi et al., 1995, Siddiqi and Titchenerhooker, 1996). Studies on the application of HPH processing to the inactivation of pathogens and spoilage organisms in foods have produced variable results. At an HPH pressure of 150 MPa, Lanciotti et al. (1996) observed variable reductions (1 to 6 log cfu/ml) in populations of E. coli, L. monocytogenes, S. aureus and Y. lipolytica in one pass through the HPH system, and noted that pre-treatment incubation temperature, pH, a.sub.w and the species of microorganism interact to affect the extent of reductions greatly. These variable results imply that the thermal history and physicochemical composition of a food can have a significant impact on the ability of HPH processing to inactivate the microbes in the food (Lanciotti et al., 1996). Using single-pass HPH of 150 MPa, Fantin et al. (1996) reduced populations of four strains of S. cerevisiae by 1 to 2 log cfu/ml and populations of four strains of Y. lipolytica by 1 to 3 log cfu/ml. Popper and Knorr (1990) observed reductions of 1 to 2 log cfu/ml in populations of B. subtilis and S. lactis subjected to one-pass HPH at 150 MPa. Bailey et al. (1995) observed that recombinant E. coli cells harvested in the stationary growth phase were more resistant to multiple-pass HPH in the range of 41 to 62 MPa than exponential growth phase cells.
It is interesting to note that researchers have noted exactly the opposite resistance trends for stationary and exponential phase cell cultures for HPH and HHP. These opposing observations for the two techniques highlights the unpredictability of these techniques for microbial inactivation and also demonstrates why they have not been implemented commercially for use as antimicrobial processes. The techniques of HPH and HHP do not provide reliable and consistent inactivation of microbes in liquids.
The U.S. Food and Drug Administration (FDA), in direct response to the Odwalla apple juice contamination, has considered mandatory pasteurization of all fresh juices, and while it decided to not require pasteurization of fresh juices at this time, the National Advisory Committee on Microbiological Criteria for Foods reported that "producers should strongly consider pasteurization until alternative risk management strategies are developed".
Thus, there is a need to develop processing alternatives. The invention described herein provides a continuous high-pressure process to treat liquids, such as milk, juices, other liquid and semi-fluid (i.e. flowable) foods, and liquid pharmaceutical preparations that destroys micro-organisms typically found in these liquids.
The invention described herein provides the advantage of being a more consistent and reliable process for inactivating microbes in fluids. This invention provides another advantage in being a continuous process, so pressure vessels of extremely large diameter are not needed. In commercial applications, continuous processes are in general more economically feasible than batch processes. Additionally, the process and apparatus of this invention allow in-line filling and sealing of containers, thus avoiding potential reintroduction of microorganisms.